Fluorescent probe and application of fluorescent detection of glyphosate

Abstract

The invention relates to the fields of chemistry, agriculture and environmental protection, discloses a fluorescent probe and application of the fluorescent probe to fluorescence detection of glyphosate, and particularly relates to rapid detection of glyphosate in a water body and agricultural products by a fluorescent probe technology. A colorimetric fluorescent probe with a 2-chloroaniline sensing unit combined with fluorophore dansyl chloride induces fluorescence enhancement through a substitution reaction with glyphosate. A fluorescence titration experiment is carried out on glyphosate with different concentrations in a range of 0-70 [mu] mol / L, the fluorescence spectrum of the probe is rapidly increased along with the gradual addition of glyphosate, and a satisfactory relation line is established between the fluorescence intensity and the concentration of glyphosate. The existence of common anions and metal ions in the water body does not interfere with the reaction, which shows that the detection method has excellent anti-interference performance. In addition, the probe can be used for detecting glyphosate in tap water, Xiangxi river reservoirs, soil, soybeans and corn, and the recovery rate range is 94.7%-109.9%. The open-type fluorescent probe can be used for specifically detecting glyphosate by naked eyes under ultraviolet light.

Description

Technical Field

[0001] The present invention relates to a technology for rapidly detecting glyphosate in water and agricultural products, and specifically to the preparation of a fluorescent probe for rapidly detecting glyphosate in water and agricultural products. Background Art

[0002] Glyphosate is a highly effective, broad-spectrum, organic phosphine herbicide launched by Monsanto in 1974. It is also the fastest growing, most widely sold, and most widely used herbicide in the world. Glyphosate is widely used in orchards, rice, corn, soybeans, wheat, tea gardens and other cash crops. Due to its high frequency of use, glyphosate has been found in surface water and streams in agriculture, cities, and forestry, posing a potential hazard to human health. It has been listed by the International Agency for Research on Cancer (IARC) as a substance with genotoxicity and potential carcinogenicity to humans, sparking intense controversy among the public and regulatory agencies.

[0003] Glyphosate has a highly polar molecular structure and lacks chromophores or fluorophores, making it difficult to detect. Conventional analytical methods for its detection include gas chromatography (GC), gas chromatography-mass spectrometry (GC-MS), high performance liquid chromatography (HPLC), enzyme-linked immunosorbent assay (ELISA), capillary electrophoresis (CE), ion chromatography (IC) and inductively coupled plasma mass spectrometry (ICP-MS). Although these methods are generally suitable for laboratory analysis with sufficient sensitivity and accuracy, they still have some inevitable disadvantages, such as time-consuming derivatization procedures (4-16 hours) and the need for well-trained personnel, which limits their practical application.

[0004] In recent years, fluorescent probes have attracted extensive attention due to their low cost and easy operation. Although a variety of fluorescent probes have been reported for the detection of glyphosate, the general design strategy is based on competitive coordination chemical sensors, Cu 2+ Added fluorescence quenching and glyphosate-induced fluorescence recovery. However, compared with the "signal on-off-on" type fluorescent probes, the on-type fluorescent probes usually have a higher signal-to-noise ratio under dark background and can effectively avoid false positive signals, which means they are more suitable for practical applications. Summary of the invention

[0005] Designing a fluorescent probe that can directly identify glyphosate is challenging. The colorimetric fluorescent probe, which combines the 2-chloroaniline sensing unit with the fluorophore dansyl chloride, induces fluorescence enhancement by undergoing a substitution reaction with glyphosate. That is, the probe shows a weak fluorescence signal. After adding glyphosate, the PET fluorescence intensity increases by more than 10 times through the photoinduced electron transfer process. This open-type fluorescent probe can achieve a stable fluorescence signal within 2 minutes. In addition, glyphosate can be specifically detected by the naked eye under ultraviolet light, showing great prospects for on-site determination.

[0006] The present invention is achieved through the following technical solutions: Fluorescent probes are used to quickly detect glyphosate in water and agricultural products. Its molecular structure is:

[0007] The synthesis method of the fluorescent probe described above:

[0008] Dissolve 2-chloroaniline and dansyl chloride in chloroform, add a little triethylamine, heat and reflux, evaporate the reaction product and purify it by silica gel column chromatography to obtain the fluorescent probe product. The reflux reaction temperature is 60-80°C and the reaction time is 10-20h.

[0009] On the one hand, triethylamine mainly acts as a base catalyst and participates in acid-base neutralization reactions; on the other hand, triethylamine can act as a proton intermediary and participate in proton transfer reactions, promote proton transfer, and accelerate the reaction rate.

[0010] The technical solution of the present invention also provides a fluorescent probe for detecting glyphosate, wherein the fluorescent probe is the fluorescent probe described above or the fluorescent probe prepared by the method described above.

[0011] The reaction principle of the present invention is: 2-chloroaniline-linked dansyl chloride is used as a fluorescent probe, which undergoes a substitution reaction with glyphosate to induce fluorescence enhancement. Its possible response mechanism is shown in the following formula:

[0012] The fluorescent probe for detecting glyphosate is used for detecting glyphosate in a non-disease diagnosis and treatment method during the glyphosate detection process.

[0013] The fluorescent probe detects glyphosate in a water environment, a solvent environment, a soil environment, and a plant environment.

[0014] The water environment includes any one of tap water, river water and reservoir; The solvent environment includes N,N-dimethylformamide, tetrahydrofuran, dimethyl sulfoxide, ethyl acetate, and acetone; The soil environment includes a soil environment to which glyphosate has been applied; The plant environment includes any one of soybean, corn, rapeseed, wheat, tea, citrus, and cotton; The present invention also provides a new use of the fluorescent probe for detecting glyphosate in selectively detecting glyphosate.

[0015] The fluorescent probe for detecting glyphosate is used for detecting glyphosate in a non-disease diagnosis and treatment method during the process of selectively detecting glyphosate.

[0016] The fluorescent probe for detecting glyphosate has a new use in selectively detecting glyphosate in a cationic environment; the cation includes Na + , K + , Cu 2+ Mg 2+ 、Zn 2+ 、Co 2+ 、Al 3+ , Fe 3+ and Cr 3+ Any one or more combinations of .

[0017] The fluorescent probe for detecting glyphosate has a new application in selectively detecting glyphosate in an anion environment; the anions include SO2-4, HCO-3, H2PO-4, CH3COO - , Cl - , NO-3, or any one or more combinations thereof.

[0018] The fluorescent probe for detecting glyphosate has a fluorescence emission peak intensity of about 530 nm during the process of selectively detecting glyphosate; The minimum detection limit was less than 4.0 nmol·L -1 The minimum detection limit is preferably less than 3 nmol·L -1 The minimum detection limit is preferably less than 2 nmol·L -1 ; The minimum detection time is less than 8 min, more preferably the minimum detection time is less than 5 min, and further preferably the minimum detection time is less than 3 min.

[0019] Advantages of the present invention: The fluorescent probe prepared by the invention has a simple synthesis process and is easy to prepare.

[0020] The fluorescent probe prepared by the present invention is effective for 0-70 μmol·L -1 The glyphosate concentration in the range was linear and the minimum detection limit was 2.0 nmol·L -1 It can meet the detection requirements of my country's drinking water hygiene standards (GB5749-2006), has high sensitivity, and realizes trace detection of glyphosate.

[0021] The fluorescent probe prepared by the invention has good anti-interference ability to glyphosate and is not interfered by other metal ions and common anions.

[0022] The fluorescent probe prepared by the present invention can detect glyphosate in tap water, Xiangxi River Reservoir, soil, soybeans and corn, with a recovery rate of 94.7-109.9%.

[0023] The fluorescent probe prepared by the present invention can identify and detect glyphosate with the "naked eye" and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is the mechanism diagram of the substitution reaction between glyphosate and the fluorescent probe.

[0025] Figure 2 For fluorescent probes 1 H NMR spectrum.

[0026] Figure 3 For fluorescent probes 13 C HMR spectrum.

[0027] Figure 4 The fluorescence spectra of the probe in response to different concentrations of glyphosate.

[0028] Figure 5 Make a graph for the glyphosate working curve.

[0029] Figure 6 is the response time of the fluorescent probe to glyphosate.

[0030] Figure 7 This is a diagram showing the effect of coexisting metal ions on glyphosate detection.

[0031] Figure 8 This is a diagram showing the effect of coexisting anions on glyphosate degradation. DETAILED DESCRIPTION

[0032] The present invention will be further described below in conjunction with specific embodiments, but the present invention is not limited to the following embodiments.

[0033] Example 1 is a method for synthesizing a fluorescent probe: 2-Chloroaniline (1.27 g, 10 mmol) and dansyl chloride (2.967 g, 11 mmol) were dissolved in chloroform (150 mL), triethylamine (1.6 mL) was added dropwise, and the mixture was refluxed at 65 °C for 12 h. After cooling, the reaction solution was concentrated by rotary evaporation, extracted with dichloromethane, and dried by adding anhydrous sodium sulfate. After rotary evaporation again, the mixture was purified by silica gel column chromatography with petroleum ether / ethyl acetate as the eluent (1:20, v / v) to obtain a light green solid with a yield of 85%. 1 H NMR, 13 C HMR spectroscopy (e.g. Figure 2 and 3 ) and high-resolution mass spectrometry analysis fully demonstrated the synthesis of the probe.1 H NMR (400 MHz, DMSO- d 6) δ 10.26 (s, 1H), 8.47(d, J = 8.0 Hz, 1H), 8.37 (d, J = 8.0 Hz, 1H), 8.03 (dd, J1= 2.0 Hz, J2= 8.0Hz, 1H), 7.59-7.54 (m, 2H), 7.32 (dd, J1= 2.0 Hz, J2= 8.0 Hz, 1H), 7.26-7.13(m, 4H), 2.82 (s, 6H). 13 C NMR (100 MHz, DMSO- d 6) δ 151.73, 136.36. 134.1,130.56, 130.32, 129.62, 129.47, 129.42, 128.32, 128.16, 127.91, 127.54,123.50, 119.66, 115.64. HR-MS (ESI): [M+H] + Calculate for C 18 H 17 ClN2O2S, 361.8573; found, 361.8570. Example 2 is the fluorescence probe for different concentrations of glyphosate (C0 = 0 ~70 μmol·L -1 ) Fluorescence response The fluorescent probe prepared in Example 1 was prepared into a 5 μmol / L stock solution using N,N-dimethylformamide (DMF) solution. 3 mL of the fluorescent probe stock solution was taken into a cuvette each time, and 0 to 70 μmol·L -1 The glyphosate standard solution was placed at room temperature for 5 minutes, and the fluorescence emission spectrum of the probe was measured under the action of 340 nm excitation light. The results are as follows Figure 4 Record the peak fluorescence intensity at 530 nm (as shown in Figure 5 ), the linear relationship between fluorescence intensity and PMG concentration was obtained as y = 800.8x + 4334.6 (R 2 =0.995).

[0034] Example 3 is the response time of the fluorescent probe and glyphosate This example uses the fluorescent probe stock solution prepared in Example 2 to detect its response time to glyphosate. Take 3 mL of the fluorescent probe stock solution in a cuvette and add 30 μmol·L -1After waiting for 0, 2, 5, and 10 minutes respectively, the fluorescence emission spectrum of the probe was measured under the action of 340nm excitation light. The results are as follows Figure 6 As shown, the fluorescence intensity hardly changed significantly after 2 minutes, indicating that the response time of the fluorescent probe after interaction with glyphosate was fast and stable.

[0035] Example 4 is the selectivity of the fluorescent probe for glyphosate in the presence of metal ions This example uses the fluorescent probe stock solution prepared in Example 2 to detect its selectivity for glyphosate. Take 3 mL of the fluorescent probe stock solution in a cuvette and add 30 μmol·L -1 glyphosate solution, and then add 10 μmol·L -1 Common metal ions (Na + , Cu 2+ Mg 2+ 、Zn 2+ 、Co 2+ 、Al 3+ , Fe 3+ and Cr 3+ In the present invention, only these common cations are used as experimental cases. The technical solutions of the present application are not limited to these metal cations. The technical solutions for detecting glyphosate in the cations are all within the protection scope of this case. Under the action of 340 nm excitation light, the fluorescence emission spectrum of the probe is measured. The results are as follows Figure 7 As shown, the fluorescence of glyphosate hardly changed significantly, indicating that the presence of metal ions would not interfere with the detection of glyphosate.

[0036] Example 5 is the selectivity of the fluorescent probe for glyphosate in the presence of anions This example uses the fluorescent probe stock solution prepared in Example 2 to detect its selectivity for glyphosate. Take 3 mL of the fluorescent probe stock solution in a cuvette and add 30 μmol·L -1 glyphosate solution, and then add 10 μmol·L -1 Common anions (SO2- 4, HCO- 3, H2PO- 4, CH3COO - , Cl - , NO- 3, the present invention only uses these common anions as experimental cases, the technical solutions of the present application are not limited to these anions, and the technical solutions for detecting glyphosate in the anions are all within the protection scope of this case), under the action of 340 nm excitation light, the fluorescence emission spectrum of the probe is measured, and the results are as follows Figure 8 As shown, the fluorescence of PMG hardly changed significantly. The results indicate that the presence of anions will not interfere with the detection of PMG. Therefore, the probe has good anti-interference ability for the detection of PMG.

[0037] Example 6: Detection of glyphosate in actual water samples using fluorescent probes In order to investigate the potential application of the probe in the actual environment, two water samples, tap water and Xiangxi River Reservoir (Yichang, Hubei), were selected and pretreated: the water samples were centrifuged at 10000 rpm for 10 min and filtered with a 0.45μ filter membrane to prepare concentrations of 10 μmol·L -1 , 30 μmol·L -1 , 50 μmol·L -1 of glyphosate solution.

[0038] The fluorescent probe stock solution prepared in Example 2 was used, and the glyphosate solutions of different concentrations were added to the fluorescent probe. Under the action of 340 nm excitation light, the peak fluorescence emission intensity of the fluorescent probe at 530 nm was measured, and the value was substituted into the equation in Example 2 to calculate the concentration of the glyphosate solution to be tested. The test results are shown in Table 1.

[0039] Example 7: Detection of glyphosate in agricultural products using fluorescent probes In order to investigate the potential application of the probe in the actual environment, campus soil, soybeans and corn (purchased from local supermarkets) were selected and the standard addition method was used for actual sample testing. Pretreatment was performed by weighing a certain amount of soybean or corn samples, grinding them, mixing them with 100 mL of water, ultrasonically treating them for 15 min, and centrifuging them at 10,000 rpm for 10 min.

[0040] The fluorescent probe stock solution prepared in Example 2 was used, and the glyphosate solutions of different concentrations were added to the fluorescent probe. Under the action of 340 nm excitation light, the peak fluorescence emission intensity of the fluorescent probe at 530 nm was measured, and the value was substituted into the equation in Example 2 to calculate the concentration of the glyphosate solution to be tested. The test results are shown in Table 1.

[0041] As can be seen from Table 1, the recovery rate of glyphosate in actual water samples is 94.7-109.9%. These results show that the fluorescent probe prepared by the present invention has high accuracy in detecting glyphosate in actual water samples and agricultural products.

[0042] Table 1 Fluorescent probes for detecting glyphosate in actual water samples and agricultural products

Claims

1. A fluorescent probe, characterized in that The structural formula of the fluorescent probe is: 。 2. The method for preparing a fluorescent probe according to claim 1, characterized in that: 2-Chloroaniline and dansyl chloride were dissolved in chloroform, triethylamine was added, and the temperature was raised to reflux for reaction. The reaction product was evaporated and purified by silica gel column chromatography to obtain a fluorescent probe product. The reaction formula is as follows: 。 3. The method for preparing a fluorescent probe according to claim 2, characterized in that: The reflux reaction temperature is 60-80°C and the reaction time is 10-20 h.

4. A fluorescent probe for detecting glyphosate, characterized in that: The fluorescent probe is the fluorescent probe described in claim 1 or a fluorescent probe prepared by the method described in claim 2 or 3.

5. The fluorescent probe for detecting glyphosate according to claim 4, characterized in that: The fluorescent probe detects glyphosate in a water environment, a solvent environment, a soil environment, and a plant environment.

6. The fluorescent probe for detecting glyphosate according to claim 5, characterized in that: The water environment includes any one of tap water, river water and reservoir; The solvent environment includes N,N-dimethylformamide, tetrahydrofuran, dimethyl sulfoxide, ethyl acetate, and acetone; The soil environment includes a soil environment to which glyphosate has been applied; The plant environment includes any one of soybean, corn, rapeseed, wheat, tea, citrus and cotton.

7. A novel use of the fluorescent probe for detecting glyphosate according to any one of claims 4 to 6 in the selective detection of glyphosate.

8. The new use according to claim 7, characterized in that: The fluorescent probe for detecting glyphosate has a new use in selectively detecting glyphosate in a cationic environment; the cation includes Na + , K + , Cu 2+ Mg 2+ 、Zn 2+ 、Co 2+ 、Al 3+ , Fe 3+ and Cr 3+ Any one or more combinations of .

9. The new use according to claim 7, characterized in that: The fluorescent probe for detecting glyphosate has a new application in selectively detecting glyphosate in an anion environment; the anions include SO2-4, HCO-3, H2PO-4, CH3COO - , Cl - , NO-3, or any one or more combinations thereof.

10. The use according to any one of claims 7 to 9, characterized in that: The fluorescent probe for detecting glyphosate has a fluorescence emission peak intensity of about 530 nm during the process of selectively detecting glyphosate; The minimum detection limit is less than 4.0 nmol·L -1 The minimum detection limit is preferably less than 3 nmol·L -1 The minimum detection limit is preferably less than 2 nmol·L -1 ; The minimum detection time is less than 8 min, more preferably the minimum detection time is less than 5 min, and further preferably the minimum detection time is less than 3 min.

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